Abstracts (31-40)

31) abstract withdrawn

32) Michael Wong, Bio 498, Poster presentation and oralAbstract Title: The Impact of Behavioural Fever on the Induction and Resolution of Inflammatory Processes

Abstract:
Fever has long been acknowledged as an important innate response to increase clearance of pathogens, but little is known about how immune function is directly modulated by this response. Ectotherms like fish can also experience a ‘behavioural’ fever, where body temperature is increased by actively seeking warmth instead of altering metabolism or intrinsic thermoregulation. To elucidate the effects of fever on the induction and resolution of inflammation, a zymosan induced behavioural fever model in goldfish (Carassius auratus) will be used in conjunction with a specially designed, ring-shaped aquarium. This design creates a temperature gradient using fluid dynamics instead of chambers, while eliminating other factors affecting behaviour.Using this model, we have demonstrated fever characterised by low velocity and high temperature preference at ~5-20h post injection. We will now examine the kinetics of pro and anti-inflammatory cytokine profiles in fever (temperature gradient) or non-fever (constant temperature) conditions throughout the 48h post-injection. Functional responses of innate immune cells like phagocytic capacity (zymosan particles, bacteria and apoptotic cells), generation of reactive oxygen intermediates, killing potential, and nuclear translocation of NFκβ will also be measured to give a multifactorial view of how fever impacts their inflammatory and immune function.

Abstract:
Transcription activator-like (TAL) effectors are DNA-binding proteins natively produced by the plant pathogen Xanthomonas to modify host gene expression. The core of these proteins is made up of repetitive elements containing 2 polymorphic amino acids. Two amino acid di-residues specify binding to each of the four DNA bases. When constructed in arrays of these repeats, the TAL effector can bind to a specific sequence of DNA. TAL effectors can be fused to the catalytic domain of the FokI nuclease to create a TAL effector nuclease (TALEN). Pairs of these constructs can then easily be designed to create double strand breaks at very specific sites in the genome. Repair of double strand breaks is an error prone process that typically results in the generation of insertion and deletion errors. This has previously been shown to be a viable method of targeted mutagenesis and is of particular interest to the zebrafish community. Within our lab, we have successfully used TALENs to create zebrafish mutants. In particular we have used this technology to create mutant models for genes implicated in eye disease. This technology has also allowed us to examine genes involved in early embryogenesis of the neural tube.

Abstract:
Restoration of native boreal forest is a required component of reclamation following oil sands mining. An integral part of the boreal ecosystem is the ectomycorrhizal (EM) fungal community which colonizes the roots of host trees. EM fungi form symbiotic relationships with most boreal trees species, providing the host with nutrients from the soil such as phosphorus and nitrogen, in exchange for carbon which is utilized for growth. My study seeks to understand community development of EM fungi within a reclamation study in the oil sands near Fort McMurray, Alberta. Research focuses on the EM fungal community 15 months after initial planting of Picea glauca, Populus tremuloides, and Pinus banksiana seedlings – trees common in the western boreal forests – in soil mediums of peat mineral mix, forest floor material, and subsoil. EM fungal diversity will be compared to a nearby reference site which has been treated with various levels of disturbance. Comparing the two sites will provide insight into EM community development at different levels of succession and site disturbance.

Abstract:
Innate immune cell effector responses, like phagocytosis and degranulation, are essential for host defense against invading pathogens. These responses are highly conserved among vertebrates, including fish, amphibians, birds and mammals. In these animals, immune cells constantly survey the extracellular and intracellular compartments for pathogens using specialized immunoregulatory receptors that can promote stimulatory or inhibitory cellular signaling events. One example of these receptor-types are the channel catfish (Ictalurus punctatus) leukocyte immune-type receptors (LITRs) that our lab uses to examine the biochemical mechanisms and functional consequences of immunoregulatory receptor-mediated signaling events. For example, IpLITR 2.6b and IpLITR 1.1b are stimulatory and inhibitory IpLITR-types that use immunoreceptor tyrosine-based activating and immunoreceptor tyrosine-based inhibitory motifs (ie. ITAMs and ITIMs), respectively, to influence immune cell functions. Previously, IpLITR2.6b was shown to promote cellular degranulation, cytokine secretion and phagocytosis which were ITAM-dependent. Conversely, IpLITR 1.1b was shown to signal through ITIM-dependent and -independent pathways resulting in the abrogation of immune cell responses including degranulation and cytokine secretion. Unexpectedly, IpLITR 1.1b was capable of inducing the phagocytic response in immune cells demonstrating that inhibitory immune receptor-types have stimulatory potential. ‘Functional plasticity’ of ITIM-encoding receptors challenges classical stimulatory/inhibitory paradigms of vertebrate receptors with the first example from Teleost reported here. To characterize IpLITR 1.1b-mediated phagocytosis, an epitope-tagged receptor was co-immunoprecipitated from stably transfected, vanadate-stimulated rat basophilic leukemia cells and membranes were blotted for candidate signaling molecules. My results have revealed that IpLITR 1.1b predictably recruits phospho-SHP2 but surprisingly also binds phospho-PI3K and phospho-Syk, which are considered stimulatory signaling molecules. This suggests that IpLITR 1.1b may employ an alternative, non-ITAM-mediated phagocytic pathway which may also represent an ancient, highly conserved signaling pathway that arose in a vertebrate ancestor.

36) Matthew Adams, Ecology/Cynthia Paszkowski, M.Sc./ oral presentationAbstract Title: The response of a population of long-toed salamanders (Ambystoma macrodactylum) in Waterton Lakes National Park to management efforts intended to reverse its decline

Abstract:
In Waterton Lakes National Park, AB, the population of long-toed salamanders inhabiting Linnet Lake has undergone a significant decline since 1994. The decline has been attributed to larval and egg depredation by fish, which invaded the system during a recent flooding event, and vehicle-caused mortality on a nearby road that salamanders cross during breeding migrations. I collected mark-recapture data in the summer and fall of 2013 to reassess the population of both salamanders and fish in Linnet Lake to see if either population has responded to treatments designed to reduce fish, and increase salamander abundance. I also used passive integrated transponder (PIT) tags and radio frequency identification (RFID) antennas to monitor salamander movement through under-road culverts to test the efficiency of motion sensing cameras currently used for this purpose. I found that the population of salamanders may have decreased slightly since a 2009 estimate and the population of fish, although lowered by fish removals in 2010 and 2011, is still large. I also found that cameras used to monitor tunnel use by salamanders were less efficient than previously estimated. These results are from the first year of a two-year project, with field work scheduled to resume in April of this year.

Abstract:
Post-glacial lakes, common feature in northern landscapes provide favorable ecosystems for studying intra-specific diversity in fishes. Great Bear Lake with its large size and virtually pristine, recently colonized cold water habitats, provides unique opportunities for Lake Trout diversification. Previous work identified three common morphotypes in the shallow-water habitat of Great Bear Lake, displaying differences in head shape and fins and a fourth rarer group. The lack of body shape variation among groups combined with the size the lake, lead us to investigate geographic morphological patterns the five arms of Great Bear Lake. Genetic and morphological distance matrices were also compared to investigate potential parallel patterns, and suggested phenotypically plastic responses to distinct environments. Inter-arm morphological variation in body shape within morphotypes reveals another layer of diversity across this large and heterogeneous northern lake. Despite increasing attention to intraspecific polymorphism in post-glacial lakes, the extent of the potential for radiation remains unknown.

Abstract:
Species detection is the first stage to creating biodiversity indices for species distributions. Amphibian monitoring by physical detection may be biased due to the secretive nature of many amphibians. This presents a need for innovative approaches to amphibian monitoring that are more sensitive and comprehensive. In aquatic habitats DNA accumulates from sloughed cellular debris. Amplifying, sequencing, and comparing short genetic fragments from the environment (environmental DNA) may identify what organisms occupy a habitat. In Alberta, Canadian toad (Anaxyrus hemiophrys) is listed as a “data deficient” species (Alberta Environment and Sustainable Resource Development 2012). We are developing an eDNA protocol to detect Canadian toads and testing it across their range in central Alberta. We selected 26 potential breeding ponds by querying Alberta’s Fisheries and Wildlife Management Information System (FWMIS) database and consultation with local researchers. Call and visual encounter surveys were conducted at each site for Canadian toad presence or absence. Four water samples were taken per site, preserved, and stored at -20°C until processed. Target loci will be amplified from samples, sequenced, and aligned to cataloged Canadian toad sequences. Results will be compared to the physical detection data to determine the reliability of the eDNA method for detecting Canadian toads.

Abstract:
Sarcopenia is the age-related loss of skeletal muscle mass and function that is attributed to fiber atrophy and loss of myofibers. The goal of this study is to identify and quantitate cell death pathways in aged Fisher 344 Brown Norway (FBN) rats. We have previously characterized sarcopenia in FBN rats and have observed an age-dependent decline in muscle mass and fiber number and an increase in fiber atrophy and collagen deposition in vastus lateralis, rectus femoris and vastus medialis muscles. We have developed a serial histological cross-sectional approach to determine myofiber cell death mechanisms in natural ageing. Our data suggest that skeletal muscle fibers in quadriceps of aged rats undergo cell death through both apoptotic and necrotic pathways. Cell death is in fibers exhibiting dysfunctional electron transport system, abnormalities that accumulate with age and result in atrophy and fiber breakage. Our data indicates that age-dependant muscle fiber loss is mediated by apoptosis and necrosis.

Abstract:
Alternative Oxidase (AOX) is a nuclear encoded (aod-1), mitochondrial protein, present in fungi, plants, as well as some animals and protists. The enzyme allows electrons to continue to flow through the first steps of the standard Election Transport Chain (sETC) when impaired in some way. AOX accepts electrons from the ubiquinone pool and passes them directly to molecular oxygen. In Neurospora crassa expression of aod-1 is very low under normal growth conditions but is induced by an unknown signal that originates from the mitochondria when the sETC is impaired. This signal is known to activate two transcription factors, AOD2 and AOD5, which are always bound as a heterodimer to the aod-1 promoter to initiate transcription. However, evidence suggests that an additional post-transcriptional mechanism of regulation exists. Mutant strain T1P11 has been shown to constitutively produce the aod-1 mRNA even though the AOX protein can be detected in the mitochondria under inducing conditions. This observation suggests N. crassa are able to regulate AOX post-transcriptionally by an undescribed mechanism. We are using qPCR and western blotting to investigate T1P11 in the hopes of better understanding this level of regulation.